Development and evaluation of a rapid CRISPR-based diagnostic for COVID-19

Hou, Tieying; Zeng, Weiqi; Yang, Minling; Chen, Wenjing; Ren, Lili; Ai, Jingwen; Wu, Ji; Liao, Yalong; Gou, Xuejing; Li, Yongjun; Wang, Xiaorui; Su, Hang; Gu, Bing; Wang, Jianwei; Teng, Xu

doi:10.1371/journal.ppat.1008705

Cited by 171 publications

(151 citation statements)

References 23 publications

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“…9,10 Today, RT-PCR and mNGS are two common molecular diagnostic approaches for 2019-nCov. 11 In RT, cDNA is synthesized from the extracted RNAs from the samples and then, in the presence of the designed and targeted primers for specific sequences (like 2019-nCov specific sequence), amplification of the designated sequence is accomplished by thermocycler. 12 In order to confirm that the amplified products are the targeted sequence, gel electrophoresis or sequencing methods could be performed.…”

Section: Pcr-based Methodsmentioning

confidence: 99%

“…thermocycler) for the isothermal molecular methods. 11 CRISPR-based method and loopmediated isothermal amplification (LAMP) are two isothermal molecular techniques.…”

Section: Pcr-based Methodsmentioning

confidence: 99%

“…Although the main aim of CRISPR is gene editing, the cleavage activities of Cas nuclease in rule-less and irregular manner results in the use of CRISPR/Cas technique in nucleic acid detection. 11 Because of the global concern about the need for specific, sensitive and rapid diagnostic techniques, tremendous attention has been attracted by CRISPR-nCov, as this technique could detect as few as 10 RNA copies/µL of 2019-nCov just within 40 minutes. 11,26 The sensitivity and speed of the technique will be increased by applying machine learning designing.…”

Section: Crispr-based Methodsmentioning

confidence: 99%

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A Review on Applicable and Available Paraclinical Methods for Diagnosis of Coronavirus Disease-19

et al. 2020

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Background: The recent outbreak by a novel coronavirus originated from Wuhan, China in 2019, and is progressively spreading to other countries. Timely diagnosis of the coronavirus disease 2019 (COVID-19) improves the survival of the patients and also prevents the transmission of the infection. In this study, we reviewed the applicable and available methods for the diagnosis of COVID-19. Methods: For the review, we systematically searched Web of Science, PubMed, and Iranian articles that were published about COVID-19 diagnostic methods with a combination of the key terms: laboratory, radiological, tests, coronavirus. Results: Although the current gold standard diagnostic test for this virus is real-time reverse-transcriptase polymerase chain reaction (RT-PCR), the occasional false-negative and the low sensitivity of the test should not be underestimated. A chest computed tomography (CT) scan is another diagnostic test for COVID-19, with higher sensitivity but low specificity. A combination of sensitive RT-PCR with a chest CT scan together with the clinical features are highly recommended for the proper diagnosis. Notably, there are some other sensitive and low-cost tests for evaluation of COVID-19 infection, but their validation should be approved. Conclusion: Since early and accurate diagnosis of the viral disease could improve the survival rate of the patients, and halt the transmission chain, it is not surprising that tremendous attempts should be made to reduce the limitations of the tests leading to the false-negative results and to find a rapid test for the diagnosis of COVID-19.

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Section: Pcr-based Methodsmentioning

confidence: 99%

“…thermocycler) for the isothermal molecular methods. 11 CRISPR-based method and loopmediated isothermal amplification (LAMP) are two isothermal molecular techniques.…”

Section: Pcr-based Methodsmentioning

confidence: 99%

Section: Crispr-based Methodsmentioning

confidence: 99%

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A Review on Applicable and Available Paraclinical Methods for Diagnosis of Coronavirus Disease-19

et al. 2020

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“…It has exhibited limit of detection in 10 fM of RNA target. This technique is much advanced than CRISPR COVID (Hou et al, 2020) as previously mentioned in this manuscript, which provides qualitative results using isothermal amplification. The sensitivity of other CRISPR based tests have been taken into consideration while developing this assay.…”

Section: Crispr-based Point Of Care Diagnosismentioning

confidence: 99%

“…Many techniques can be exploited to make rapid and affordable testing available to common people that include Point of Care (POC) also known as bed side testing. POC testing is developing as a portable and promising tool in pathogen diagnosis (Yeh et al, 2017), a major example is the discovery of CRISPR COVID that has been shown to deliver comparable specificity and sensitivity as that of RT-PCR and sequencing based meta genomic diagnostic approaches (Hou et al, 2020). Moreover, lateral flow assay (LFA) is one of the latest POC techniques, which works on the principle of fluid flow, and it is employed in immune assays targeting protein, DNA or RNA based samples (Tang et al, 2017).…”

Section: Introductionmentioning

confidence: 99%

An Update on Molecular Diagnostics for COVID-19

Islam

Iqbal

2020

Front. Cell. Infect. Microbiol.

101

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SARS‐CoV‐2 RNA Detection by a Cellphone‐Based Amplification‐Free System with CRISPR/CAS‐Dependent Enzymatic (CASCADE) Assay

Silva

Erdogmus

Shokr

et al. 2021

Adv Materials Technologies

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CRISPR (Clustered regularly interspaced short palindromic repeats)‐based diagnostic technologies have emerged as a promising alternative to accelerate delivery of SARS‐CoV‐2 molecular detection at the point of need. However, efficient translation of CRISPR‐diagnostic technologies to field application is still hampered by dependence on target amplification and by reliance on fluorescence‐based results readout. Herein, an amplification‐free CRISPR/Cas12a‐based diagnostic technology for SARS‐CoV‐2 RNA detection is presented using a smartphone camera for results readout. This method, termed C ellphone‐based a mplification‐free s ystem with C RISPR/C A S‐ d ependent e nzymatic (CASCADE) assay, relies on mobile phone imaging of a catalase‐generated gas bubble signal within a microfluidic channel and does not require any external hardware optical attachments. Upon specific detection of a SARS‐CoV‐2 reverse‐transcribed DNA/RNA heteroduplex target (orf1ab) by the ribonucleoprotein complex, the transcleavage collateral activity of the Cas12a protein on a Catalase:ssDNA probe triggers the bubble signal on the system. High analytical sensitivity in signal detection without previous target amplification (down to 50 copies µL −1 ) is observed in spiked samples, in ≈71 min from sample input to results readout. With the aid of a smartphone vision tool, high accuracy (AUC = 1.0; CI: 0.715 – 1.00) is achieved when the CASCADE system is tested with nasopharyngeal swab samples of PCR‐positive COVID‐19 patients.

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Development and evaluation of a rapid CRISPR-based diagnostic for COVID-19

Cited by 171 publications

References 23 publications

A Review on Applicable and Available Paraclinical Methods for Diagnosis of Coronavirus Disease-19

A Review on Applicable and Available Paraclinical Methods for Diagnosis of Coronavirus Disease-19

An Update on Molecular Diagnostics for COVID-19

SARS‐CoV‐2 RNA Detection by a Cellphone‐Based Amplification‐Free System with CRISPR/CAS‐Dependent Enzymatic (CASCADE) Assay

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